P53和P21在PC12细胞分化中的作用

Roles of P53 and P21 in the differentiation of PC12 cells

目的探讨P53及其下游P21蛋白在PC12细胞分化中的可能作用机制。方法用反转录病毒质粒pBabe-P53/m175转染未分化PC12细胞,经筛选后建立野生型P53蛋白功能丧失的细胞系PC12(P53/m175);利用倒置相差显微镜、流式细胞术及Western blot等方法,比较两种细胞在神经生长因子(NGF)作用后,细胞分化表型、细胞周期及相关蛋白表达的改变。结果NGF作用于正常PC12细胞组,P53/P21蛋白表达持续升高,细胞G1期阻滞出现的时间与蛋白表达开始增加的时间相一致;而在NGF作用的PC12(P53/m175)细胞组,细胞不能表达P21蛋白,且细胞周期G1期阻滞的程度也出现显著下降。NGF作用下,两组细胞都能观察到神经突起的生长,表现出明显的分化表型。结论PC12细胞经NGF作用后出现P53及P21蛋白持续表达增加,主要介导了细胞分化过程中细胞周期G1期阻滞这一特征的出现,但并不是导致神经突起进行性生长这一特征的必需条件。

Objective To investigate the roles of P53/P21 during neuronal differentiation with a differentiated model of PC12 cells. Methods A new cell line PC12（P53/m175） was created by stable transfection of a retrovirus plasmid pBabe-P53/m175, which contains a dominant-negative P53 gene mutant. After NGF treatment, observing with phase-contrast microscopy, flow cytometric analysis and western blotting of P53 and P21 were performed. Results Expression of P53 and P21 was obviously increased in NGF-induced PC12 cells. The appearance of cell cycle G1 phase arrest paralleled the increased expressions of P53 and P21. The level of P21 protein did not change after treatment with NGF in PC12（P53/m175） cells and the extent of G1 phase arrest markedly decreased. However, we did find the normal neurite outgrowth in NGF-treated PC12 （P53/m175） cells. Conclusion NGF-induced an increased protein levels of P53 and its transcriptional element, P21 is essential for cell cycle G1 phase arrest, but does not necessarily correlate with the neurite outgrowth of PC12 cells.